1. Field of the Invention
The invention relates to a microfluidic device. More particularly, the invention relates to a non-transparent microfluidic device with surface plasmon resonance (“SPR”) capabilities.
2. Description of the Prior Art
The use of conventional SPR as a testing tool offers several advantages. For example, it is relatively fast, it does not require labeling and it can be performed on site. Further, SPR provides for good sensitivity and can be performed in “real time,” which generates kinetic information, i.e., one can see the binding while it occurs as opposed to after rinsing off the reagents.
SPR is an optical phenomenon which occurs as a result of total internal reflection of light at a metal film-liquid interface. Total internal reflection is observed in situations where light travels through a medium such as glass, and is reflected back through that medium from the interface with a different medium, for example a liquid buffer solution. In order for total internal reflection to occur, the angle of incidence of the light must be greater than a critical angle determined by the refractive indices of the optical media. Although the light is totally reflected, a component of the incident light momentum, termed the evanescent wave or surface plasmon, penetrates a distance, on the order of one wavelength, into the medium, e.g., the buffer, on the opposite side of the interface.
If the incident light is monochromatic and polarized, and the interface between the media is coated with a thin metal film, such as gold or silver, having a thickness which is a fraction of the wavelength of the incident light, the evanescent wave can interact with free oscillating electrons, or plasmons, in the metal film surface. The plasmons will absorb energy from the evanescent wave at a particular angle of incidence, which is dependent upon the refractive index of the liquid medium adjacent to the metal film, i.e. within a distance of about 300 nm from the metal film. Thus, for a given refractive index in the liquid, the intensity of the reflected light varies according to the angle of incidence of the light, and there is a sharp drop in the intensity of the reflected light at a particular angle at which peak absorbance occurs. This angle can be termed the “resonance angle.”
Changes in the refractive index of a buffer solution, for example, will alter the resonance angle. By measuring the angle at which the peak occurs, it is possible to detect changes in the refractive index of the buffer solution. Because proteins, for example, in the buffer solution alter its refractive index, it is possible to measure, and monitor continuously, the protein content in the buffer solution adjacent to the metal film by measuring the resonance angle. Further, the interaction of macromolecules in the buffer solution with, for example, surface immobilized ligand or non-specific binding polymers, e.g., antibody binding to peptide or protein, causes a change in the refractive index. This change results in a correlative change in the resonance angle, which is detectable and quantifiable.
SPR technology is utilized in commercially available instruments, for example, produced by BIACORE (Uppsala, Sweden). Further, a SPR detector is manufactured by Pharmacia Biosensor AB (Uppsala, Sweden).
The BIACORE methodology relies on immobilization of ligands onto the surface of a sensor chip consisting of a glass substrate having a gold film covered by a monolayer of a long hydroxyalkyl thiol to which is covalently attached a thin layer of carboxymethylated dextran. The immobilization procedure is performed with the sensor chip in place in the instrument and is continuously monitored by the SPR detector.
The sensor chip is contacted by a microfluidic cartridge which has formed on it a number of channels which define the flow of samples across the surface of the sensor chip. The microfluidic cartridge, which is in place when the ligand is introduced to the sensor, contains pneumatic valves, which control the flow of samples through the channels.
An unknown sample or ligate solution is introduced into the apparatus to contact the immobilized ligand. The interaction between ligand and ligate is observed directly by surface plasmon resonance techniques and the measurements recorded on a computer via a program such as Bialogue, produced by Pharmacia BioSensor AB. The sensor chip is discarded to waste after interacting with the ligand, however, the microfluidic cartridge is reusable.
The BIACORE system is limited to a glass microfluidic sensor chip given that a transparent material is necessary to allow visible light produced by the BIACORE optical detector to reach the metal film on the sensor chip.
As indicated above, SPR has traditionally been performed with a glass substrate or microfluidic chip. The inventors of the present invention have discovered that SPR may be effectively performed on non-transparent substrates or chips, which offer a number of advantages over glass. Given the above detailed utility of SPR sensing, the expansion of SPR sensing to microfluidic devices made from materials other than transparent glass is highly significant.
Silicon based sensors for measuring radiation reflected off the surface plasmon generating film on a glass substrate are used in some existing SPR systems. Given the heightened sensitivity of silicon sensors at 800 nm, the optical detectors of some SPR systems are configured to generate 800 nm radiation so as to take advantage of this heightened sensor sensitivity. Accordingly, SPR systems, which produce non-visible radiation, i.e., operating in the infra-red region, are known. To date, however, the inventor is not aware of any such SPR system used in combination with a substrate or microfluidic chip made from non-transparent material.
While the prior art microfluidic systems may be suitable for the particular purpose employed, or for general use, they are not as suitable for the purposes of the present invention as disclosed hereafter.